PROJECT SUMMARY Traumatic brain injuries (TBI) affect ~1.6 to 3.8 million athletes annually, most commonly in the form of concussion, leaving 10-40% of patients with long-lasting (>1 month) physical, emotional, psychological and cognitive symptoms. Moreover, repetitive head impacts sustained within the brain's ?window of vulnerability? post-injury can lead to cumulative severity of these adverse consequences, which can last longer than the effects of a single insult alone and can potentially cause permanent degenerative conditions. Objective biomarkers that identify those at highest risk of long-lasting symptoms and/or that identify the safe return-to- play interval could be an important component of individualized patient care. To address this unmet clinical need, we obtained preliminary data in mice that suggests low cerebral blood flow measured at baseline or early on in the evolution of injury caused by repetitive mild closed head injury could be a useful predictor of ultimate cognitive outcome. This proposal will build upon our existing preclinical data to further explore the utility of blood flow, as well as several other promising biomarkers of cognitive outcome, including cerebral oxygen metabolism, cerebrovascular reactivity, and resting-state functional connectivity. Because of innate limitations of clinical experiments of mTBI, including patient/injury heterogeneity and time of assessment, we will perform initial testing of these biomarkers in a closed head mouse model of sports-related head injury that features blunt impact followed by rotational head acceleration. First, we will thoroughly quantify how blood flow, oxygen metabolism, vascular reactivity, and functional connectivity change with time before, during, and after repetitive closed head injury (Aim 1). We will quantify these parameters using three non-invasive optical modalities (frequency-domain near-infrared spectroscopy, diffuse correlation spectroscopy, and optical intrinsic signal imaging of functional connectivity) that are well-suited for longitudinal monitoring. From these data, we will identify several critical time points in order to investigate the prognostic ability of blood flow, oxygen metabolism, vascular reactivity, and functional connectivity to identify mice that will develop worse cognitive outcomes (Aim 2a). Outcome will be quantified with a battery of tests for spatial learning and memory, anxiety, and depression. Finally, we will test whether cerebral blood flow and/or oxygen metabolism can be used as markers of the brain's vulnerable window following mTBI (Aim 2b). We will test whether mice whose blood flow and/or oxygen metabolism levels return to baseline prior to subsequent closed head injury have improved outcomes compared to those who do not return to baseline before subsequent closed head injury. Upon completion of the proposed aims, we will not only be poised to translate these biomarkers to well-controlled clinical experiments, but also to embark on future studies that delineate molecular mechanisms of blood flow, oxygen metabolism, vascular reactivity, and functional connectivity derangements in the context of novel therapies.